Ribosome dysfunction underlies SLFN14-related thrombocytopenia.

Pathogenic missense variants in SLFN14, which encodes an RNA endoribonuclease protein that regulates rRNA degradation, are known to cause inherited thrombocytopenia with impaired platelet aggregation and ATP secretion. Despite rather mild laboratory defects, these patients display an obvious bleeding phenotype. The function of SLFN14 in megakaryocyte (MK) and platelet biology is unknown. This study aims to model the disease in the immortalized megakaryocyte cell line imMKCL and characterize the platelet transcriptome in patients with a SLFN14 K219N variant. MK derived from heterozygous and homozygous SLFN14 K219N imMKCL and stem cells of blood from patients mainly presented with a defect in proplatelet formation and mitochondrial organization. SLFN14 defective platelets and mature MK showed signs of rRNA degradation while this was absent in undifferentiated imMKCL cells and granulocytes. Total platelet RNA was sequenced for two patients and 19 healthy controls. Differential gene expression analysis yielded a total of 2999 and 2888 significantly (|log2FC|>1, FDR<0.05) up- and downregulated genes, respectively. Remarkably, these downregulated genes were not enriched for any biological pathway while upregulated genes were enriched for pathways involved in (mitochondrial) translation and transcription with a significant upregulation of 134 ribosomal protein genes (RPG). Upregulation of (M)RPG through increased mTORC1 signaling in SLFN14 K219N MK seems to be a compensatory response to rRNA degradation. Indeed, mTORC1 inhibition with rapamycin resulted in further enhanced rRNA degradation in SLFN14 K219N MK. Taken together, our study indicates dysregulation of mTORC1 coordinated ribosomal biogenesis as the disease mechanism for SLFN14-related thrombocytopenia.